1. Field of the Invention
The present invention relates to a method for producing nucleotides by fermentation. Nucleotides such as nucleoside 5′-phosphate esters are useful as seasonings, drugs, raw materials thereof and so forth.
2. Description of the Related Art
As methods for industrial production of nucleoside 5′-phosphate esters, there are known methods comprising producing nucleoside by fermentation and enzymatically phosphorylating the obtained nucleoside to obtain nucleoside 5′-phosphate ester.
On the other hand, methods of directly producing nucleoside 5′-phosphate esters by fermentation have also been proposed. For example, Japanese Patent Publication (Kokoku) No. 56-12438 discloses a method for producing 5′-guanylic acid, which comprises culturing a mutant strain of a bacterium belonging to the genus Bacillus showing adenine auxotrophy and resistance to decoyinine or methionine sulfoxide and having an ability to produce 5′-guanylic acid (guanosine 5′-monophosphate, also abbreviated as “GMP” hereinafter) and collecting GMP produced and accumulated in the medium. Further, there are several reports on deriving strains which produce 5′-inosinic acid (inosine 5′-monophosphate, also abbreviated as “IMP” hereinafter) from inosine producing strains of Bacillus subtilis (Magasanik, B. et al., J. Biol. Chem., 226, 339 (1957); Fujimoto, M., et al., Agr. Biol. Chem., 30, 605 (1966)). However, the production of nucleoside 5′-phosphate esters by direct fermentation generally suffers from insufficient yield, and it is not so practical compared with the aforementioned enzymatic methods.
As the reasons for the difficulty of IMP production by direct fermentation, there are mentioned bad cell permeability of IMP and quite ubiquitous distribution of degradative enzymes that decompose IMP (Nucleic Acid Fermentation, Edited by Aminosan Kakusan Shudankai, Kodansha Scientific, Japan). To overcome these obstacles, there has been attempted to delete nucleotide degradative activity. As degradative enzymes that decompose IMP into inosine, 5′-nucleotidase, acid phosphatase, alkaline phosphatase and so forth are conceived (Nucleic Acid Fermentation, supra). Further, the aforementioned Japanese Patent Publication No. 56-12438 also suggests that a bacterial strain showing high GMP yield can be obtained from a mutant strain showing reduced nucleotidase activity.
As a technique for producing nucleoside 5′-phosphate ester on an industrial level, a method of producing IMP by using a mutant strain of Brevibacterium ammoniagenes has been developed (Furuya et al., Appl. Microbiol., 16, 981 (1968)).
As described above, various studies have been made on the production of nucleoside 5′-phosphate esters by direct fermentation, and some successful examples are also known. However, there are many unknown points about nucleotide degradative enzymes, and it cannot be said that improvement of yield has been studied sufficiently. In particular, no example of production of nucleoside 5′-phosphate esters on a practical level has been known for bacteria belonging to the genus Escherichia. 